The oligodendroglial reaction to brain stab wounds: An immunohistochemical study

Summary Myelin/oligodendrocyte specific protein was compared to glial fibrillary acidic protein and 23-cyclic nucleotide 3-phosphodiesterase expression in normal rat brains and following stab wounds to the cerebral cortex, corpus callosum and hippocampus. Animals with stab wounds were allowed to recover for 5,15,28, 45 and 70 days post-operation before fixation by perfusion. Sections were reacted with antibodies against myelin/oligodendrocyte specific protein, glial fibrillary acidic protein and 23-cyclic nucleotide 3-phosphodiesterase, and observed by light and electron microscopy. Normal cerebral cortex had very few myelin/oligodendrocyte specific protein-positive and 23-cyclic nucleotide 3-phosphodiesterasepositive cells, but some glial fibrillary acidic protein-positive cells. The myelinated fibres of the corpus callosum were heavily stained for myelin/oligodendrocyte specific protein but unstained by glial fibrillary acidic protein or 23-cyclic nucleotide 3-phosphodiesterase antibodies. Some immunopositive cells were present in the corpus callosum and hippocampus with all three antibodies. After stab wound myelin/oligodendrocyte specific protein-positive reactive cells had more and longer processes and stained more intensely than equivalent cells in normal brain. These cells were distributed along the wound track, including within the cerebral cortex. The numbers of these cells increased until 28 days post-operation and then decreased so that very few were found at 70 days post-operation except in the corpus callosum. Where demyelination occurred myelin/oligodendrocyte specific protein-staining was lost. Staining for 23-cyclic nucleotide 3-phosphodiesterase revealed a similar pattern. Glial fibrillary acidic protein-positive reactive cells, which were also more robust than the normal cells, were more widely distributed. They increased in number throughout the time periods studied and gliosis was evident on the contralateral side. The glial fibrillary acidic protein-positive astrocytes were also different from the myelin/oligodendrocyte specific protein-positive and 23-cyclic nucleotide 3-phosphodiesterase-positive oligodendrocytes in terms of cell shape. With electron microscopy myelin/oligodendrocyte specific protein-positive cells showed features typical of immature oligodendrocytes. We conclude that the injury caused a numerical increase in oligodendrocytes and that myelin/ oligodendrocyte specific protein is a good marker for the oligodendroglial response and demyelination in pathological conditions.     

Dissociation of acetylcholine- and cyclic GMP-induced currents inXenopu oocytes

InXenopus follicular oocytes, activation of muscarinic receptors evokes a slow potassium current (H-response); a similar current is evoked by intracellular injection of cyclic guanosine 3,5-monophosphate, cGMP (Dascal et al. 1984). We have tested the hypothesis that cGMP may be the second messenger that mediates the opening of K channel by acetylcholine (ACh). ACh elevated the intracellular level of cGMP with a time course similar to that of the development of the muscarinic H-response; maximal increase in cGMP concentration above the control was about 0.2 pmole/oocyte. The amount of injected cGMP that produced a detectable K current (threshold dose) varied between 0.5 and 3 pmole/oocyte. At low doses of cGMP, the slope of log dose-log response curve was about 2.5, suggesting involvement of a biochemical process with a positive cooperativity of at least 3. Higher doses of cGMP evoked, in addition to the outward current, an irregular, rapidly developing, long-lasting inward current, that never reached amplitudes comparable to those of ACh-evoked Cl currents. The K current elicited by cGMP was insensitive to elevation or depletion of external Ca. It was potentiated by isobutylmethylxanthine (IBMX). ACh strongly inhibited the cGMP-evoked K current when applied at the plateau of the latter. 4-Phorbol 12,13-dibutyrate (PDBu) (1 M) rapidly and completely inhibited the cGMP response. It is concluded, that most of the results presented in this report contradict the hyothesis that cGMP is the intracellular mediator of ACh-induced changes in membrane conductance in the oocytes.Abbreviations ACh acetylcholine - cAMP cyclic adenosine 3,5-monophosphate - cGMP cyclic guanosine 3,5-monophosphate - EGTA ethylenediaminetetraacetic acid - Hepes N-2-hydroxyethyl-piperazinc-N-2-hydroxypropanesulphonic acid - IBMX 3-isobutyl-l-methylxanthine - IP₃ inositol 1,4,5-trisphosphate - PDBu 4-phorbol 12,13-dibutyrate     

Cyclische Nucleotide als intracelluläre Überträgerstoffe bei Hormonwirkungen

Zusammenfassung Hormone dienen als extracelluläre Informationsüberträger zwischen ihrem Bildungsort, einer endokrinen Drüse, und den Zellen, deren Funktion sie regulieren. Durch die Reaktion des Hormons mit den an der Zellmembran gelegenen Receptoren wird die Aktivität der mit diesen eng verknüpften Adenyl-Cyclase beeinflußt. Die meisten Hormone erhöhen in ihrem Zielorgan die Aktivität dieses Enzyms und führen hierdurch zu einem raschen Anstieg der intracellulären Konzentration von Adenosin-3:5-monophosphat (Ado-3:5-P). Dieses cyclische Nucleotid wird durch eine spezifische Phosphodiesterase zu Adenosin-5-monophosphat abgebaut. Auch die Aktivität dieses Enzyms bestimmt die intracelluläre Ado-3:5-P-Konzentration, die im Vergleich zu der anderer Nucleotide sehr gering ist.Ado-3:5-P beeinflußt als zweiter, intracellulärer Überträgerstoff die Aktivität zahlreicher Schlüsselenzyme. Die Ado-3:5-P-Konzentration bestimmt hierdurch das Gleichgewicht verschiedener Stoffwechselwege zueinander und damit die Reaktion einer Zelle auf eine hormonale Stimulierung. An einer Reihe von Enzymen wird die durch Ado-3:5-P bedingte Aktivitäts-Änderung durch einen gleichartigen Mechanismus bewirkt. Das cyclische Nucleotid stimuliert Proteinkinasen, die eine Phosphatgruppe des ATP auf verschiedene Proteine übertragen und hierdurch deren Eigenschaften verändern können. So steigt bei Phosphorylierung durch eine Ado-3:5-P-stimulierbare Proteinkinase die Aktivität der Triglyceridlipase und der Glykogen-Phosphorylase-b-kinase an, dagegen nimmt die Aktivität der Glykogen-Synthetase ab; durch Phosphorylierung von Histonen kann deren Repressorcigenschaft vermindert und die Synthese von Enzymen gesteigert werden.In manchen tierischen Geweben wurde auch eine spezifisch durch Guanosin-3:5-monophosphat (Guo-3:5-P) stimulierbare Proteinkinase nachgewiesen. Dieses cyclische Nucleotid kommt wie Ado-3:5-P in allen Säugerorganen vor. Die Bildung von Guo-3:5-P aus GTP wird durch die Guanyl-Cyclase katalysiert, ein Ferment, das im Gegensatz zur Adenyl-Cyclase zum großen Teil nicht an die Zellmembranen gebunden ist. Die Konzentration von Guo-3:5-P in verschiedenen Geweben, im Blutplasma und im Urin wird durch Hormone beeinflußt. Es ist noch nicht bekannt, welche hormonalen Regulationen durch Guo-3:5-P vermittelt werden; dagegen ist bei vielen, rasch einsetzenden Hormonwirkungen die Beteiligung von Ado-3:5-P nachgewiesen worden.

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Abkürzungen Ado-3:5-P Adenosin-3:5-monophosphat - dAdo-3:5-P Desoxy-adenosin-3:5-monophosphat - Guo-3:5-P Guanosin-3 : 5-monophosphate - Nuc-3:5-P Nucleosid-3:5-monophosphat - NTP Nucleosidtriphosphat - NMP Nuclcosid-5-monophosphat - dATP Desoxyadenosintriphosphat - P_i anorganisches Phosphat - PP_i anorganisches Pyrophosphat - DNS Desoxyribonucleinsäure - RNS Ribonucleinsäure - r-RNS ribosomale RNS - m-RNS Boten-RNS - Glykogen-Synthetase UDP-Glucose--1,4-glucan--4-glucosyltransferase - ICSH interstitial-cell-stimulating hormone     

Studien über den Mechanismus der Immunhämolyse: Der Bindungsmodus der vierten Komplementkomponente

Zusammenfassung Die Bindung der vierten Komplementkomponente (C₄) an sensibilisierte Hammelblutzellen (EA) ist von der vorhergehenden Bindung der ersten Komplementkomponente abhängig. Das Präparat EAC₁ kann, wenn ihm C₄ in geeigneter Form angeboten wird, in den Komplex EAC_1,4 überführt werden. Hierbei verschwindet der titrierbare Gehalt der flüssigen Phase an C₄; gleichzeitig acquirieren die Zellen die Fähigkeit, mit R₄ zu lysieren. Der zeitliche Ablauf dieser Veränderung wird untersucht. Die Eeaktion zwischen EAC₁ und C₄ verläuft äußerst schnell, und ihre Geschwindigkeit wird durch die Reaktionstemperatur nicht beeinflußt. Während die Bindung von C₁ an EA nur in Gegenwart von Ca⁺⁺ erfolgt, läuft die Bindung von C₄ an EAC₄ auch in Abwesenheit von zweiwertigen Metallionen ab. Die vonLevine u.Mayer als ein Ganzes betrachtete Ca⁺⁺-abhängige Überführung von EA in EAC_1,4 besteht mithin aus zwei aufeinanderfolgenden Teilreaktionen, von denen die erste (C₁-Bindung) calciumabhängig und die zweite (C₄-Bindung) von zweiwertigen Ionen unabhängig ist.Mit Unterstützung der Deutschen Forschungsgemeinschaft sowie der Gesellschaft der Freunde und Förderer der Medizinischen Akademie Düsseldorf.     

Determination of the 5′ and 3′ terminal noncoding sequences of the bi-segmented genome of the avibirnavirus infectious bursal disease virus

Summary Terminal sequences of the bi-segmented dsRNA genome of 3 different strains of infectious bursal disease virus (IBDV) were analyzed by the rapid amplification of cDNA 5 ends (5RACE) procedure. Both segments are 85% homologous in a 32-nucleotide sequence comprising the 5 end, whereas the 3 end has a conserved pentamer. Comparison to published terminal sequences of other IBDV strains revealed high conservation between the two segments but more serotype-specific nucleotide changes (5 on segment A and 3 on segment B) in the 5 noncoding region compared to the 3 noncoding region (none on segment A and 1 on segment B).     

Inverse Correlation Between IgG-Antihinge Region and Antierythrocyte Autoantibody in Chronic Benign and Malignant Cold Agglutination

Previous reports provided evidence of an immunosuppressive role of natural anti-F(ab)₂ antibodies. If suppressive anti-F(ab)₂ antibodies also regulated the autoantibody production in cold agglutination, one would expect high titers of anti-F(ab)₂ to be associated with low titers of cold agglutinins. Indeed, our previous studies revealed an inverse correlation between IgG-anti-F(ab)₂ and cold agglutinins. Many previous experiments focused on anti-F(ab)₂ of an antiidiotypic nature. Recent epitope mapping showed that anti-F(ab)₂ of healthy persons is not an antiidiotype but recognizes a hinge region sequence. We attempted to answer the question whether this IgG-antihinge antibody is responsible for the previously described association between anti-F(ab)₂ and cold agglutinins. IgG-antihinge and IgG-anti-F(ab)₂ antibody was determined and statistically analyzed in the serum of 334 patients with cold agglutination. Our experiments revealed a strong correlation between the concentrations of antihinge and the previously described anti-F(ab)₂ antibody. The anti-F(ab)₂ activity was competitively inhibited by a synthetic hinge peptide. Moreover, patients with high antihinge titers had low cold agglutinin titers, and vice versa. A stratification according to cold agglutinin specificity and disease etiology showed that the inverse correlation is present only in anti-I and anti-i patients suffering from monoclonal B-lymphocyte proliferation. In conclusion, our results confirm the correlation previously described for anti-F(ab)₂ antibody and antierythrocyte autoantibody and define for the first time an association between an idiotype-independent anti-IgG autoantibody and cold agglutinin.     

Molecular Analysis of Fiji Disease Fijivirus Genome Segments 1 and 3

Fiji disease fijivirus (FDV) genomic segments 1 (S1) and 3 (S3) were completely sequenced. FDV S1 comprised 4532nt and was predicted to encode a 170.6kDa protein. FDV S3 comprised 3623nt and was predicted to encode a 135.5kDa protein. The terminal sequences of S1 and S3 were 5 AAGUUUUU......CAGCUAGCGUC 3 and 5 AAGUUUUU......CAGCAGAUGUC 3, respectively, and located immediately adjacent to these sequences were 12bp imperfect inverted repeats. The predicted translation product of FDV S1 showed highest similarity to Rice black-streaked dwarf virus (RBSDV) S1 and is thought to encode the viral RNA-dependent RNA polymerase (RdRp). The predicted translation product of FDV S3 was found to be most similar to RBSDV S4 which is thought to encode the 'B-spike' protein. The FDV sequence contained an ATP/GTP binding motif and a leucine zipper motif, but these motifs were not found in the RBSDV sequence. Phylogenetic analysis based on the amino acid sequences of the RdRp of FDV S1 and other reoviruses revealed that the fijiviruses form a cluster close to the oryzaviruses. The RdRp sequences were grouped into genera that were consistent with the current reovirus classification scheme that is based on physico-chemical and biological properties.     

Regulation of the trehalose-6-phosphate synthase complex in Saccharomyces

Summary Trehalose-6-phosphate synthase is another example of an enzyme of carbohydrate metabolism, in Saccharomyces, which could be regulated by interconversion of forms. Deactivation was mediated both in vivo and in vitro by a cyclic AMP-dependent protein kinase. Reversibility of this process was obtained by a phosphatase treatment leading to an increase in activity. The phosphorylated, less active form of the enzyme proved to be more susceptible to activation by ATP.Mg. Mutants with well defined lesions in the cyclic AMP-dependent protein kinase system were used to corroborate our findings of a possible regulatory mechanism of trehalose-6-phosphate synthase activity by interconversion of forms.Abbreviations PMSF phenyl-methyl-sulfonyl fluoride - G-6-P glucose-6-phosphate - UDPG uridine-5-diphosphoglucose - PEP phosphoenol pyruvate - NAD⁺ -nicotinamine adenine dinueleotide - ATP adenonise 5-triphosphate - cAMP adenosine 2:3-cyclic monophosphate - MOPS 3 (N-morpholino) propanesulfonic acid     

Experimentelle Pyelonephritis

Zusammenfassung Gesamtlytische Serumkomplement (C)-Aktivität sowie Antikörper(Ak)-Titer wurden zu verschiedenen Zeiten bei experimenteller nichtobstruktiver Enterokokken-Pyelonephritis untersucht. In der akuten Infektionsperiode ergibt sich nach anfänglichem C-Titerabfall ein ausgeprägter Anstieg der antibakteriellen Serumantikörper mit dann entgegengesetztem Verhalten der C-Titerkurve. Bei fortgeschrittener chronischer Pyelonephritis sind bei weiterhin erhöhten Ak-Titern unauffällige C-Titer nachzuweisen.Die C-Titerhöhe ergibt damit keine diagnostischen oder prognostischen Hinweise für das entzündliche Geschehen einer chronischen Pyelonephritis.

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Summary Total lytic C activity and antibody titer in experimental non-obstructive enterococcal pyelonephritis was measured at different times. In the period of acute infection an initial C-titer decrease with a definite rise of antibacterial serum antibodies is found with a subsequent increase to near normal levels of the C-titer curve. In progressive chronic pyelonephritis increased bacterial antibody titers with normal C-titers are found. — The height of the C-titer level therefore does not give any diagnostic or prognostic leads about the inflammatory state of a chronic pyelonephritis.

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Die vorliegenden Untersuchungen wurden mit Hilfe der Deutschen Forschungsgemeinschaft durchgeführt.

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Herrn Prof. Dr.R. Brühl, Trier, zum 70. Geburtstag gewidmet.     

Proteins are attached to the ends of a linear plasmid in the filamentous fungus Ascobolus immersus

Summary In seven out of eleven wild strains of the Ascomycete Ascobolus immersus plasmid DNA was found. There was great variability with respect to size and number of the plasmids in the strains concerned. For a further analysis two plasmids originating from one wild strain were submitted to restriction analysis and electron microscopy. Both turned out to be linear having different molecular weights (pAIl = 7.9 kb, pAI2 = 5.6 kb). Denaturation of pAI2 and subsequent renaturation revealed the presence of inverted repeats (0.7 kb) at both ends. After treatment with proteinase K and 5 and 3 specific exonucleases it became evident that the 5 ends of pAI2 are linked with proteins. In this respect it is similar in structure to other linear genetic elements such as the linear plasmids found in Zea mays and the genomes of adenoviruses.Dedicated to Prof. Dr. Fritz Kaudewitz on the occasion of his 65th birthday     

Identification of an ADPG-dependent trehalose synthase in Saccharomyces

Summary Uridine diphosphoglucose is not the sole donor for trehalose synthesis in yest cells: an ADPG-dependent trehalose synthase, has been identified in mutant strains with undetectable UDPG-dependent trehalose-6-P synthase activity. Genetic and chromatographic studies indicate that the two activities correspond to different proteins. The apparent K K_m for the nucleotide is similar for both enzymes, and Mg²⁺ is also required for both activities; however, a striking difference was observed with respect to ATP.Mg activation. This newly determined enzymatic activity in Saccharomyces clarifies previous contradictory results with mutant strains that are able to accumulate trehalose during growth yet whose UDPG-dependent trehalose synthase activity is undetectable in vitro.Abbreviations PMSF Phenyl-methyl-sulfonyl fluoride - EDTA ethileno-daminotetracetic acid - G6P glucose-6-phosphate - UDPG uridine5-diphosphoglucose - ADPG adenosine-5-diphosphoglucose - UDP uridine-5-diphosphate - ADP adenosine-5-diphosphate - PEP phosphoenol pyruvate - ATP adenosine-5-triphosphate - UTP uridine-5triphosphate - Pi inorganic phosphate - MOPS 3 (N-morpholino) propanesulfonic acid - PNPG paranitrophenylglucoside     

Genome Organization and Expression of the Penicillium stoloniferum Virus S

The complete sequences of two double-stranded RNAs (dsRNAs) (referred to S1 and S2) of Penicillium stoloniferum virus S (PsV-S) were established. The S1 dsRNA was 1,690bp in length, and it contained a unique open reading frame (ORF) of 539 amino acids (molecular weight of 62kDa, referred to P62). The S2 dsRNA was 1,523bp in length, and also it contained one ORF of 434 amino acids (molecular weight of 47kDa, referred to P47). Both S1 and S2 ORFs were identified only on the positive strand of each dsRNA segment. A sequence motif of (5-CUG-3) was found at the 3-termini of the positive strands of PsV-S1 and S2 dsRNAs. The predicted amino acid sequences of S1 dsRNA showed high sequence homology with the putative RNA-dependent RNA polymerases of RNA viruses. Near full-length and positive-sense single-stranded RNAs derived from the S1 and S2 dsRNAs were detected from the PsV-infected host cell. The expressed proteins of P62 and P47 showed a positive reaction against PsV-S antiserum in Western blot analysis. Phylogenetic analysis using the RDRP sequences and the capsid proteins of the various partitiviruses revealed that PsV-S is a definite member of the partitivirus, the family Partitiviridae, and especially clusters well along with D. destructiva virus 1 and 2.     

Nucleotide sequence and determination of the extremities of the 26S ribosomal RNA gene in wheat mitochondria: evidence for sequence rearrangements in the ribosomal genes of higher plants

Summary The nucleotide sequence of the wheat mitochondrial 26S ribosomal RNA gene and flanking regions was determined and compared with mitochondrial 26S rRNA genes from maize and Oenothera. All three genes exhibit a high degree of homology except within two variable regions. When the plant mitochondrial 26S rRNA genes are compared with Escherichia coli 23S rRNA and chloroplast 23S and 4.5S rRNA genes, a third variable region is apparent close to the 3 end of the gene. The 5 and 3 ends of the wheat mitochondrial gene were determined by S1 nuclease mapping. Computer analysis of the wheat mitochondrial gene revealed several small sequences present either in the 5 region of the 26S rRNA gene or in the 18S rRNA gene.     

Rhinovirus detection using probes from the 5′ and 3′ end of the genome

Summary This study investigated the abilities of cDNA probes from the 5 and 3 ends of the genome of human rhinoviruses (HRV-) 14, 9, and 1B to detect RNA from 59 rhinovirus serotypes. The results show that probes from the 5 end of the genomes of HRV-14, 9, and 1B detected a large number of serotypes but the detection rate was variable and depended on the degree of homology with the particular probe. In contrast, all the 3 end probes were specific for the homologous virus. However, along HRV-9 probe detected a large number of serotypes.It was concluded that such cDNA probes would not detect all serotypes with equal efficiency. Synthetic oligonucleotides corresponding to short but highly conserved regions in the 5 non coding region may overcome this problem.     

Selection and characterization of pyrG mutants of Penicillium chrysogenum lacking orotidine-5′-phosphate decarboxylase and complementation by the pyr4 gene of Neurospora crassaa

Summary Pyrimidine auxotrophs of Penicillium chrysogenum have been isolated at a high frequency among mutants resistant to 5-fluoroorotic acid (5.2 mM). Some of the pyrimidine auxotrophs (e.g. strain pyrG1) showed no reversion. A radiometric assay based on the conversion of (6-¹⁴C)orotidine 5-monophosphate (OMP) into (6-¹⁴C)uridine 5-monophosphate (UMP) was developed to determine OMP-decarboxylase activity. One of the pyrimidine auxotrophs (P. chrysogenum pyrGl) was studied in detail. It was deficient in OMP-decarboxylase activity, whereas the parental strain (P. chrysogenum Wis. 54-1255) showed a normal enzyme activity. A five-fold higher OMP-decarboxylase activity was found in a P. chrysogenum pyrGI clone transformed with plasmids containing the Neurospora crassa pyr4 gene (which codes for the same enzyme).Abbreviations OMP orotidine 5-monophosphate - UMP uridine 5-monophosphate     

Effects of hydration and dehydration on cyclic adenosine 3′,5′-monophosphate concentration in the rat kidney

Summary Renal 3,5-AMP concentration has been measured in two groups of rats, one of which was hydrated, the other one was dehydrated by withdrawal of drinking-water for a period of 4–5 days. The first group was assumed to have low, the latter to have high vasopressin (ADH) plasma concentrations.The 3,5-AMP concentration in the kidney of hydrated rats was 1.7 nmoles/g (w.w.). Studies on the distribution of 3,5-AMP between various regions of the kidney revealed that the 3,5-AMP content of the cortex was higher than in the outer medulla. 3,5-AMP concentration in the inner medulla was below the sensitivity of the method used for the determination of the nucleotide.Dehydration led to a doubling of renal 3,5-AMP concentration. The increment in the cortex was greater than in the outer medulla. In the inner medulla 3,5-AMP concentration remained below the sensitivity of the method used.Renal 3,5-AMP phosphodiesterase has been obtained from two subcellular fractions, one fraction bound to large particles, probably cell membranes, the other one soluble in the supernatant. Vasopressin did alter the activity of the enzyme in neither fractions. From this it has been concluded that the increase in 3,5-AMP concentration in the kidney of dehydrated rats results from stimulation of adenyl cyclase by ADH.This study was supported by the Deutsche Forschungsgemeinschaft.Deceased October 31, 1967.     

Stereochemical aspects of the interaction of myosin and actomyosin with nucleotides

Summary The five possible analogues of ATP and the three possible analogues of ADP which contain single non-bridging sulphur atoms instead of oxygen in the polyphosphate structure have been used as probes of the interaction of nucleotides with myosin and actomyosin. Evidence is presented for the requirement of an , , -tridentate complex of magnesium and ATP as the substrate for myosin. Of the four possible tridentate MgATP diastereomers, the exo isomer (nomenclature of Cornelius & Cleland, 1978) appears to be the actual substrate.Abbreviations ATP Adenosine-5-O-triphosphate - ATP(-S) Adenosine-5-O-(1-thiotriphosphate) - ATP(-S) Adenosine-5-O-(2-thiotriphosphate) - ATP(-S) Adenosine-5-O-(3-thiotriphosphate) - ADP Adenosine-5-O-diphosphate - ADP(-S) Adenosine-5-O-(1-thiodiphosphate) - ADP(-S) Adenosine-5-O-(2-thiodiphosphate) - Enzyme Myosin ATPase (EC 3.6.1.3)     

A structural model for the genome of echovirus 22

Summary We have proposed previously that the structural model for the echovirus 22 genome is a single-stranded RNA molecule that has folded back upon itself to form a stable hairpin at the 5-terminus. The vRNA of echovirus 22 has been characterized further by digestion with selective ribonucleases, electrophoresis in composite gels, hydrodynamic studies in density gradients of Cs₂SO₄ and sucrose, thermal denaturation and 3-terminal ribonucleotide analysis. Based on these observations, the genome of echovirus 22 is a single-stranded RNA molecule having a region of secondary structure located at the 5-terminus that may be characterized as a snapback hairpin with hydrogen-bonded base-pairing. In addition, a VPg-like protein is attached (presumably to the 5-end of the RNA) and the 3-terminus contains a polyadenylic acid tract [poly(A)].     

Translation enhancer in the 3′-untranslated region of rotavirus gene 6 mRNA promotes expression of the major capsid protein VP6

Summary. The eleven rotavirus mRNAs contain 5-cap structures and most end with the 3-consensus sequence 5-UGACC-3. The UGACC functions as a common translation enhancer (3-TE-con) that upregulates viral protein expression through a process mediated by the nonstructural protein NSP3. To address the possibility that gene-specific enhancers are also contained in the untranslated regions (UTRs) of the rotavirus mRNAs, we used rabbit reticulocyte lysates to investigate the translation efficiencies of analog RNAs containing viral-specific 5-and 3-UTRs and the open reading frame for chloramphenicol acetyltransferase. These experiments combined with the analysis of full-length viral RNAs and RNAs containing 3-truncations showed that a highly active enhancer was present near the 5-end of the 139-nucleotide 3-UTR of the gene 6 mRNA (3-TEg6). The 3-TEg6 represents a functionally independent enhancer, as no other portion of the gene 6 mRNA was required for its activity. The 3-TEg6 differs significantly from the 3-TE-con in that the gene 6-specific enhancer does not require viral protein for activity and is formed by a sequence unique to only one of the eleven viral mRNAs. Together, our findings suggest that the 3-UTR of the gene 6 mRNA contains two TEs, one is gene-specific (3-TEg6) and the other is common to nearly all rotavirus genes (3-TE-con). The activity of the 3-TEg6 is likely important for directing the efficient translation of the gene 6 mRNA at levels sufficient to provide the 780 copies of VP6 necessary for the assembly of each progeny virion.     

Specificity repertoire of lymphocytes from multiple myeloma patients. I. High frequency of B cells specific for idiotypic and F(ab′)2-region determinants on immunoglobulin

The specificity repertoire of B lymphocytes from 14 multiple myeloma patients has been studied using the technique of Epstein-Barr virus (EBV) transformation of peripheral blood lymphocytes (PBL) coupled with clonal analysis by limiting dilution. We find that up to 100% of the B cells from myeloma patients undergoing EBV transformation secrete IgM specific for determinants on the F(ab)₂ region of autologous and/or heterologous monoclonal immunoglobulin. In normal individuals 0.02–0.73% of the transformed B cells secrete IgM specific for F(ab)₂ determinants. Two patients with monoclonal gammopathy of undetermined significance had only a weak reactivity to F(ab)₂ fragments. The number of anti-F(ab)₂ B cells was up to 145-fold greater in patients than in normal donors. The majority of antibodies from patient clones recognized determinants shared among 3–12 different F(ab)₂ fragments, whereas those originating from normal donor B cells saw determinants expressed on only one or two of the panel of test F(ab)₂ fragments. There was a preference for autologous M components and a high proportion of antiidiotypic reactivity in five of eight patients so analyzed. We speculate that these findings indicate the existence of an anti-F(ab)₂ immunoregulatory network mediating patient immunodeficiency, thereby creating an abnormality that may enable the progression of multiple myeloma.